Yves Théorêt

Pharmacogenomics and metabolite measurement for 6-mercaptopurine therapy in inflammatory bowel disease

BACKGROUND & AIMS The effects of 6-mercaptopurine (6-MP) are mediated via its intracellular conversion to 6-thioguanine (6-TG) and 6-methylmercaptopurine (6-MMP) nucleotide metabolites, the latter genetically controlled by thiopurine methyltransferase (TPMT). We sought to determine optimal therapeutic 6-MP metabolite levels and their correlation with medication-induced toxicity and TPMT genotype. METHODS Therapeutic response was determined in 92 pediatric patients with inflammatory bowel disease coincidentally with hematologic, pancreatic, and hepatic laboratory parameters, and compared with erythrocyte metabolite levels and TPMT genotype. RESULTS Clinical response was highly correlated with 6-TG levels (P < 0.0001) but not with any other variable, including 6-MMP levels, drug dose, gender, and concurrent medications. The frequency of therapeutic response increased at 6-TG levels > 235 pmol/8 x 10(8) erythrocytes (P < 0.001). Hepatotoxicity correlated with elevated 6-MMP levels (>5700 pmol/8 x 10(8) erythrocytes; P < 0.05). Although leukopenia was associated with higher 6-TG levels (P < 0.03), it was observed in only 8% of responders. Patients heterozygous for TPMT (8/92) had higher 6-TG levels (P < 0.0001), and all responded to therapy. CONCLUSIONS 6-MP metabolite levels and TPMT genotyping may assist clinicians in optimizing therapeutic response to 6-MP and identifying individuals at increased risk for drug-induced toxicity.

Use of alternative and complementary therapies in children with cancer

The use of complementary and alternative medicines (CAM) is becoming increasingly popular. Although considered beneficial by users, the potential for interaction or substitution with conventional treatment should not be overlooked by health care professionals. It is therefore important to gain insight into the prevalence and the factors related to the use of CAM. To establish the prevalence of use of CAM among children with cancer treated in a large pediatric hospital, describe the profile of use and factors related with use. As a secondary objective we aimed at measuring quality of life of the children aged 5 or more and compare the scores between users and non‐users.

Pharmacokinetics of Proton Pump Inhibitors in Children

The use of proton pump inhibitors (PPIs) has become widespread in children and infants for the management of paediatric acid-related disease. Pharmacokinetic profiles of only omeprazole and lansoprazole have been well characterised in children over 2 years of age with acid-related diseases. Few data have been recently published regarding the pharmacokinetics of pantoprazole in children, and none are available for rabeprazole or esomeprazole. The metabolism of PPI enantiomers has never been studied in the paediatric population.A one-compartment model best describes the pharmacokinetic behaviour of omeprazole, lansoprazole and pantoprazole in children, with important interindividual variability for each pharmacokinetic parameter. Like adults, PPIs are rapidly absorbed in children following oral administration; the mean time to reach maximum plasma concentration varies from 1 to 3 hours. Since these agents are acid labile, their oral formulations consist of capsules containing enteric-coated granules. No liquid formulation is available for any of the PPIs. Thus, for those patients unable to swallow capsules, extemporaneous liquid preparations for omeprazole and lansoprazole have been reported; however, neither the absolute nor the relative bioavailabilities of these oral formulations have been studied in children. Intravenous formulations are available for omeprazole (in Europe), lansoprazole and pantoprazole.PPIs are rapidly metabolised in children, with short elimination half-lives of around 1 hour, similar to that reported for adults. All PPIs are extensively metabolised by the liver, primarily by cytochrome P450 (CYP) isoforms CYP2C19 and CYP3A4, to inactive metabolites, with little unchanged drug excreted in the urine. Similar to that seen in adults, the absolute bioavailability of omeprazole increases with repeated dosing in children; this phenomenon is thought to be due to a combination of decreased first-pass elimination and reduced systemic clearance. The apparent clearance (CL/F) of omeprazole, lansoprazole and pantoprazole appears to be faster for children than for adults. A higher metabolic capacity in children as well as differences in the extent of PPI bioavailability are most likely responsible for this finding. This may partly account for the need in children for variable and sometimes considerably greater doses of PPIs, on a per kilogram basis, than for adults to achieve similar plasma concentrations. Furthermore, no studies have been able to demonstrate a statistically significant correlation between age and pharmacokinetic parameters among children. Despite the small number of very young infants studied, there is some evidence for reduced PPI metabolism in newborns. The limited paediatric data regarding the impact of CYP2C19 genetic polymorphism on PPI metabolism are similar to those reported for adults, with poor metabolisers having 6- to 10-fold higher area under the concentration-time curve values compared with extensive metabolisers.Finally, because a pharmacokinetic/pharmacodynamic relationship exists for PPIs, the significant interindividual variability in their disposition may partly explain the wide range of therapeutic doses used in children. Further studies are needed to better define the pharmacokinetics of PPIs in children <2 years of age.

Influence of GST gene polymorphisms on busulfan pharmacokinetics in children.

Busulfan (BU) is a key compound in conditioning myeloablative regimens for children undergoing hematopoietic stem cell transplantation (HSCT). There are wide interindividual differences in BU pharmacokinetics, which increase the risk of veno-occlusive disease, graft rejection and disease relapse. As BU is mainly metabolized by glutathione S-transferase (GST), it is hypothesized that functional polymorphisms in GST genes may explain in part the variability in BU pharmacokinetics. We analyzed polymorphisms in GSTA1 (C-69T, A-513G, G-631T, C-1142G), GSTM1 (deletion) and GSTP1 (A1578G, C2293T) genes in 28 children undergoing HSCT. All patients had individualized dosing based on pharmacokinetics after the first dose of intravenous BU. GSTM1-null individuals had higher drug exposure (PCmax=0.008; PAUC=0.003; PCss=0.02) and lower clearance (PCL=0.001). Multivariate regression models showed that, other than the drug dose and age, the GSTM1 genotype was the best predictor of first-dose pharmacokinetic variability. GSTM1-null patients also received lower cumulative BU doses (P=0.02). No association was found between BU exposure and major GSTA1 or GSTP1 gene variants. In children, GSTM1 polymorphism seems to modify BU pharmacokinetics after intravenous drug administration.

High-performance liquid chromatography-electrochemical detection of vecuronium and its metabolites in human plasma.

A high-performance liquid chromatographic assay coupled with electrochemical detection has been developed for the determination of vecuronium and its three putative deacetylated metabolites in human plasma. A novel solid-phase extraction procedure allowed good recovery of both vecuronium and its metabolites, together with ease and speed of execution. This method was sensitive, reproducible and accurate over the therapeutic range of concentrations of vecuronium and its metabolites, and was applied successfully to a study of the pharmacokinetics of vecuronium in anaesthetized patients.

Therapeutic drug monitoring for triazoles: A needs assessment review and recommendations from a Canadian perspective

Therapeutic drug monitoring (TDM) is necessary for certain drugs to ensure that the levels are sufficient to be effective, but not so high as to cause adverse effects. This review summarizes the literature regarding TDM for newer-generation extended-spectrum triazoles, including when TDM may be necessary for each drug and why, and laboratory techniques used for the measurement of levels of these drugs. The document includes recommendations for the use of TDM for each triazole that is discussed.

Association between busulfan exposure and outcome in children receiving intravenous busulfan before hematopoietic stem cell transplantation

Background and Objective: Intravenous (IV) busulfan (Bu) combined with therapeutic drug monitoring-guided dosing is associated with better event-free survival (EFS), lower transplant-related mortality. But optimal target steady-state concentration (Css) of Bu in children undergoing hematopoietic stem cell transplantation (HSCT) remains unclear. This study aimed to evaluate the relation between Css of Bu and clinical outcomes in children receiving Bu before HSCT. Methods: This study includes 75 children receiving IV Bu in 16 doses, with first dose assigned based on age. Bu first-dose pharmacokinetic parameters were estimated from Bu plasma concentrations measured at 6 time points by high-performance liquid chromatography. Doses were adjusted at the fifth dose to a target Css of 600–900 ng/mL. Cumulative incidence of overall survival (OS), EFS, transplant-related mortality, acute graft-versus host disease (aGVHD), and other toxicities in relation to Css of Bu were analyzed using Kaplan–Meier curves in univariate and Coxs proportional hazards model in multivariate analysis. Results: After the first dose, median Css was 578 (325–1227) ng/mL. Forty-one patients had Bu IV dose increased by > 10%. Neutrophil and platelet recoveries, grade 2–4 aGVHD, and nonrelapse mortality (NRM) incidences were 90%, 91%, 12%, and 13%, respectively. Relapse incidence was 33%. Incidence of veno-occlusive disease, hemorrhagic cystitis, and lung toxicities were 13%, 24%, and 7%, respectively. OS and EFS were 70% and 58%. First-dose Bu Css >600 ng/mL was associated with a higher NRM (P < 0.001) and grade 2–4 aGVHD (P = 0.04), a lower EFS (P < 0.001), and OS (P = 0.001). Conclusions: This study demonstrated a significant association between the first-dose pharmacokinetics of Bu and NRM, OS, and EFS. Bu therapeutic drug monitoring provides information that potentially influences outcomes of HSCT in pediatric patients.

A simplified method for busulfan monitoring using dried blood spot in combination with liquid chromatography/tandem mass spectrometry

RATIONALE Busulfan (Bu) is an important component of the myeloablative conditioning regimen prior to hematopoietic stem cell transplantation (HSCT) especially in children. Intravenously administered Bu exhibits a therapeutic window phenomenon requiring therapeutic drug monitoring. Analytical methods developed for Bu routine monitoring were aimed at using low volumes of biological fluids and development of simple procedures to facilitate the dosage adjustment. In this report, we describe a simple, rapid method for Bu measurement using dried blood spots (DBS) from only 5 μL of whole blood. METHODS Bu extracted from DBS with methanol was measured by high-performance liquid chromatography with electrospray ionization and tandem mass spectrometry in multiple reaction monitoring mode using D8-Bu as an internal standard. The method was in-house validated evaluating trueness, repeatability, within-laboratory reproducibility, specificity and the lower limit of quantification (LLOQ). RESULTS The method was linear in the calibration range of 100-2000 ng mL(-1) (r(2)>0.99) encompassing the therapeutic concentrations of Bu. A good trueness (<14%), precision (<10%), and recovery (100%) were observed during validation of the method with quality controls of 300, 600 and 1400 ng mL(-1). The LLOQ was determined as 50 ng mL(-1) and no matrix or carryover effects were observed. The validated method was applied to measure Bu levels in four children receiving infusion of Bu prior to HSCT. A good correlation was observed between the Bu levels measured by DBS and dried plasma spot (DPS) (r(2) =0.96) and between DPS and the GC/MS method (r(2) =0.92). Bu was found to be stable in DBS up to 6 h at room temperature and for 24 h at 4 °C. CONCLUSIONS The new DBS method facilitates earlier dosage adjustment during Bu therapy by its specific and simple procedure using 5 μL of whole blood.

Association of busulfan exposure with survival and toxicity after haemopoietic cell transplantation in children and young adults: a multicentre, retrospective cohort analysis

BACKGROUND Intravenous busulfan combined with therapeutic drug monitoring to guide dosing improves outcomes after allogeneic haemopoietic cell transplantation (HCT). The best method to estimate busulfan exposure and optimum exposure in children or young adults remains unclear. We therefore assessed three approaches to estimate intravenous busulfan exposure (expressed as cumulative area under the curve [AUC]) and associated busulfan AUC with clinical outcomes in children or young adults undergoing allogeneic HCT. METHODS In this retrospective analysis, patients from 15 centres in the Netherlands, USA, Canada, Switzerland, UK, Italy, Germany, and Australia who received a busulfan-based conditioning regimen between March 18, 2001, and Feb 12, 2015, were included. Cumulative AUC was calculated by numerical integration using non-linear mixed effect modelling (AUCNONMEM), non-compartmental analysis (AUC from 0 to infinity [AUC0-∞] and to the next dose [AUC0-τ]), and by individual centres using various approaches (AUCcentre). The main outcome of interest was event-free survival. Other outcomes of interest were graft failure or relapse, or both; transplantation-related mortality; acute toxicity (veno-occlusive disease or acute graft versus-host disease [GvHD]); chronic GvHD; overall survival; and chronic-GvHD-free event-free survival. We used propensity-score-adjusted Cox proportional hazard models, Weibull models, and Fine-Gray competing risk regressions for statistical analyses. FINDINGS 790 patients were enrolled, 674 of whom were included: 274 (41%) with malignant and 400 (59%) with non-malignant disease. Median age was 4·5 years (IQR 1·4-10·7). The median busulfan AUCNONMEM was 74·4 mg × h/L (95% CI 31·1-104·6), which correlated with the standardised method AUC0-∞ (r2=0·74), but the latter correlated poorly with AUCcentre (r2=0·35). Estimated 2-year event-free survival was 69·7% (95% CI 66·2-73·0). Event-free survival at 2 years was 77·0% (95% CI 72·1-82·9) in the 257 patients with an optimum intravenous busulfan AUC of 78-101 mg × h/L compared with 66·1% (60·9-71·4) in the 235 patients at the low historical target of 58-86 mg × h/L and 49·5% (29·2-66·0) in the 44 patients with a high (>101 mg × h/L) busulfan AUC (p=0·011). Compared with the low AUC group, graft failure or relapse occurred less frequently in the optimum AUC group (hazard ratio [HR] 0·57, 95% CI 0·39-0·84; p=0·0041). Acute toxicity (HR 1·69, 1·12-2·57; p=0·013) and transplantation-related mortality (2·99, 1·82-4·92; p<0·0001) were significantly higher in the high AUC group (>101 mg × h/L) than in the low AUC group (<78 mg × h/L), independent of indication; no difference was noted between AUC groups for chronic GvHD (<78 mg × h/L vs ≥78 mg × h/L, HR 1·30, 95% CI 0·73-2·33; p=0·37). INTERPRETATION Improved clinical outcomes are likely to be achieved by targeting the busulfan AUC to 78-101 mg × h/L using a new validated pharmacokinetic model for all indications. FUNDING Research Allocation Program and the UCSF Helen Friller Family Comprehensive Cancer Center and the Mt Zion Health Fund of the University of California, San Francisco.

Glutathione S-transferase gene variations influence BU pharmacokinetics and outcome of hematopoietic SCT in pediatric patients

BU is a key compound of conditioning regimens in children undergoing hematopoietic SCT (HSCT). Inter-individual differences in BU pharmacokinetics (PKs) might affect BU efficacy and toxicity. As BU is mainly metabolized by glutathione S-transferase (GST), we investigated the relationship between GSTA1, GSTM1 and GSTP1 genotypes with first-dose BU PKs, and the relationship with HSCT outcomes in 69 children receiving myeloablative conditioning regimen. GSTM1 null genotype correlated with higher BU exposure and lower clearance in patients older than 4 years (P⩽0.04). In accordance with the suggested functional role, GSTA1*A2 haplotype was associated with lower drug levels and higher drug clearance (P⩽0.03). Gene-dosage effect was also observed (P⩽0.007). GSTA1 haplotypes were associated with HSCT outcomes. Patients with two copies of haplotype *A2 had better event free survival (P=0.03). In contrast, homozygous individuals for haplotypes *B and *B1 had higher occurrence of veno-occlusive disease (P=0.009). GSTM1 null individuals older than 4 years had more frequently graft versus host disease (P=0.03). In conclusion, we showed that GST gene variants influence BU PK and outcomes of HSCT in children. A model for the dosage adjustment with the inclusion of genetic and non-genetic factors should be evaluated in a future prospective validation cohort.